Image-guided, spatially localized 31P magnetic resonance spectroscopy (MRS) was used to study in vivo murine cardiac metabolism under resting and dobutamine-induced stress conditions. Intravenous dobutamine infusion (24 mug. min-1. kg body wt-1) increased the mean heart rate by approximately 39% from 482 +/- 46 per min at baseline to 669 +/- 77 per min in adult mice. The myocardial phosphocreatine (PCr)-to-ATP (PCr/ATP) ratio remained unchanged at 2.1 +/- 0.5 during dobutamine stress, compared with baseline conditions. Therefore, we conclude that a significant increase in heart rate does not result in a decline in the in vivo murine cardiac PCr/ATP ratio. These observations in very small mammals, viz., mice, at extremely high heart rates are consistent with studies in large animals demonstrating that global levels of high-energy phosphate metabolites do not regulate in vivo myocardial metabolism during physiologically relevant increases in cardiac work.